Project Summary/Abstract O-linked N-acetyglucosaminylation (O-GlcNAcylation) is a reversible posttranslational modification that plays a key role in ionizing radiation (IR)-induced DNA damage response. O-GlcNAcylation is catalyzed by O- GlcNActransferase (OGT), which transfers N-acetyl-D-glucosamine from UDP-GlcNAc to serine or threonine residues of proteins. This posttranslational modification is also removed by O-GlcNAcase (OGA). In our studies, we have shown that O-GlcNAcylation is significantly enriched at DNA lesions. Since the acceptors of O-GlcNAcylation are serine or threonine residues, O-GlcNAcylation competes with DNA damage-induced phosphorylation, which in turn regulates DNA damage repair. Thus, we hypothesize that O-GlcNAcylation is a key molecular event in response to IR treatment, and targeting O-GlcNAcylation can be an effective therapeutic strategy to cancer treatment. In this research proposal, we plan to focus on one major O-GlcNAcylation substrate MDC1, and examine the role of O-GlcNAcylated MDC1 in DNA damage response including the phosphorylation events on MDC1, MDC1 governed protein ubiquitination cascade, and DNA double-strand break repair. Using O-GlcNAcylated MDC1 as the readouts, we will analyze the biological functions of both OGT and OGA in IR-induced DNA damage repair, and explore the inhibition of OGA as a novel therapeutic strategy to treat BRCA1 or BRCA2- deficient tumors in vivo.